Base-pair Ambiguity and the Kinetics of RNA Folding

Abstract

Background: A folding RNA molecule encounters multiple opportunities to form non-native yet energetically favorable pairings of nucleotide sequences. Given this forbidding free-energy landscape, mechanisms have evolved that contribute to a directed and efficient folding process, including catalytic proteins and error-detecting chaperones. Among structural RNA molecules we make a distinction between "bound" molecules, which are active as part of ribonucleoprotein (RNP) complexes, and "unbound," with physiological functions performed without necessarily being bound in RNP complexes. We hypothesized that unbound molecules, lacking the partnering structure of a protein, would be more vulnerable than bound molecules to kinetic traps that compete with native stem structures. We defined an "ambiguity index"-a normalized function of the primary and secondary structure of an individual molecule that measures the number of kinetic traps available to nucleotide sequences that are paired in the native structure, presuming that unbound molecules would have lower indexes. The ambiguity index depends on the purported secondary structure, and was computed under both the comparative ("gold standard") and an equilibrium-based prediction which approximates the minimum free energy (MFE) structure. Arguing that kinetically accessible metastable structures might be more biologically relevant than thermodynamic equilibrium structures, we also hypothesized that MFE-derived ambiguities would be less effective in separating bound and unbound molecules. Results:We have introduced an intuitive and easily computed function of primary and secondary structures that measures the availability of complementary sequences that could disrupt the formation of native stems on a given molecule-an ambiguity index. Using comparative secondary structures, the ambiguity index is systematically smaller among unbound than bound molecules, as expected. Furthermore, the effect is lost when the presumably more accurate comparative structure is replaced instead by the MFE structure. Conclusions:A statistical analysis of the relationship between the primary and secondary structures of non-coding RNA molecules suggests that stem-disrupting kinetic traps are substantially less prevalent in molecules not participating in RNP complexes. In that this distinction is apparent under the comparative but not the MFE secondary structure, the results highlight a possible deficiency in structure predictions when based upon assumptions of thermodynamic equilibrium.

Intrinsic Regulatory Role of RNA Structural Arrangement in Alternative Splicing Control

“…Considering given opportunities to form many energetically favored RNA structures exclusively to the native one, albeit at the expense of high energy- and time-consuming processes of RNA structural rearrangements, it is profound the way cells employ and manage factors introducing effective kinetics of functional RNA folding [ 70 ]. Therefore, there has been a continual necessity to confront collected knowledge from in vitro studies with a highly complex environment of living cells which provide a great range of additional factors like compartmentalization, concentration of biomolecules and protein-protein interaction network contributing to RNA conformation.…”